Activity-dependent glassy cell mechanics â : Mechanical properties measured with active microrheology.

Active microrheology was conducted in living cells by applying an optical-trapping force to vigorously fluctuating tracer beads with feedback-tracking technology. The complex shear modulus G(ω)=G'(ω)-iG″(ω) was measured in HeLa cells in an epithelial-like confluent monolayer. We found that G(ω)∝(-iω)1/2 over a wide range of frequencies (1 Hz < ω/2π < 10 kHz). Actin disruption and cell-cycle progression from G1 to S and G2 phases only had a limited effect on G(ω) in living cells. On the other hand, G(ω) was found to be dependent on cell metabolism; ATP-depleted cells showed an increased elastic modulus G'(ω) at low frequencies, giving rise to a constant plateau such that G(ω)=G0+A(-iω)1/2. Both the plateau and the additional frequency dependency ∝(-iω)1/2 of ATP-depleted cells are consistent with a rheological response typical of colloidal jamming. On the other hand, the plateau G0 disappeared in ordinary metabolically active cells, implying that living cells fluidize their internal states such that they approach the critical jamming point.

[Case of Unresectable Gastric Mixed Neuroendocrine Neoplasm(MiNEN)Treated with Chemotherapy].

A 68-year-old man diagnosed with gastric mixed neuroendocrine-non-neuroendocrine neoplasia(MiNEN)concomitant with liver metastasis received chemotherapy using ramucirumab and paclitaxel. A decrease in tumor marker levels and size of the metastatic liver lesions was observed after 3 courses of treatment. However, the patient developed progressive disease after 9 courses of chemotherapy; hence, nivolumab chemotherapy was initiated. Although liver metastases were reduced after 2 courses of nivolumab, the patient developed new liver lesions after 18 courses of treatment; irinotecan, S-1 and oxaliplatin, and trifluridine/tipiracil were then administered. Liver metastases progressed despite changing the regimen, and the patient died 25 months after the initiation of chemotherapy. Gastric MiNEN usually shows poor prognosis, and there is lack of consensus regarding optimal treatment. Ramucirumab and nivolumab are relatively well-tolerated and may be effective for chemotherapy.

Assessment of intrathoracic lymph nodes by FDG PET/CT in patients with asbestos-related lung cancer.

BACKGROUND: This study explored the assessment of intrathoracic lymph node metastasis by 18F-fluoro-2-deoxy-D-glucose (FDG) positron emission tomography computed tomography (PET/CT) in patients with asbestos-related lung cancer (ARLC). METHODS: We retrospectively reviewed the data on 35 patients with ARLC who underwent preoperative FDG-PET/CT and surgical resection between January 2012 and December 2018. We collected medical information from medical records and imaging systems and examined the FDG uptake in each lymph nodal region resected by surgery and the presence or absence of pathological lymph node metastasis. RESULTS: Pathological lymph node metastases were detected in 14 (8.70%) of 161 nodal stations. The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of FDG-PET/CT were 71.4% (10/14), 87.8% (129/147), 35.7% (10/28), 97.0% (129/133), and 86.3% (139/161), respectively. Six of the eight false-positive patients had bilateral accumulations, whereas all six true-positive patients had unilateral accumulation (P=0.006). On histopathological examination, the false-positive nodes showed disruption of lymphoid follicles in the cortex, infiltration of histiocyte-like cells in the medulla, fibrous micronodules, and severe anthracosis. CONCLUSIONS: PET/CT scans of patients with ARLC showed comparable sensitivity and specificity to those of PET/CT scans of patients with conventional lung cancer reported in the literature. Many false-positive cases also showed bilateral symmetric accumulation. This method can be used to evaluate lymph node involvement in lung cancer.

Primary omental torsion diagnosed and treated laparoscopically: a case report.

Omental torsion, a rare cause of acute abdomen in children and adults, is difficult to correctly diagnose before surgery because it mimics the common causes of acute surgical abdomen. We present a case of greater omental torsion that was diagnosed by laparoscopy. A 37-year-old man presented with right lower abdominal pain and was suspected to have appendicitis. Blood tests revealed elevated C-reactive protein and white blood cell count, whereas computed tomography revealed a nodular mass and high-density lesions in the fat tissue. As the patient's abdominal symptoms were severe and a clear diagnosis could not be made, we performed exploratory laparoscopy. Laparoscopy revealed omental torsion, and an omentectomy was performed. The patient's pain had significantly reduced post-surgery, and post-operative recovery was uneventful. Thus, laparoscopic examination is useful for accurately diagnosing omental torsion and is less invasive than surgery.

Catamenial pneumothorax due to heterotopic endometriosis in the pericardium.

A 46-year-old woman presented with a right pneumothorax at a regular medical examination during menstruation. The pneumothorax resolved without intervention; however, 6 months later, she was referred to our hospital due to chest pain and dyspnoea. A chest X-ray showed grade III pneumothorax and surgery was performed. During surgery, the patient was found to have pleural adhesions around the right upper lung, pores in the diaphragm and a blueberry spot in the pericardium. The margins of the upper lobe and diaphragm were covered with a polyglycolic acid sheet at the operation. Eight days after surgery, she was referred to our hospital again due to massive haemothorax. The reoperation suggested that the aforementioned blueberry spot in the pericardium was the source of bleeding. The spot was resected and shown to be oestrogen and progesterone receptor-positive, providing evidence of heterotopic endometriosis.

[A Case of Intraductal Apocrine Carcinoma of the Breast].

A female in her 60s who complained of nipple discharge in her left breast for 1 year. A soft mass ill-defined margin in the border of AB area was observed. Mammography showed a focal asymmetric density. Ultrasonography disclosed an irregular heterogenous low echoic lesion in the AB area of her left breast. MRI image showed an enhanced lesion in the inner area. The pathological diagnosis by core needle biopsy was non-invasive ductal carcinoma with apocrine metaplasia. Mastectomy with sentinel lymph node biopsy of the left breast was performed. Post operative histopathological examination revealed intraductal apocrine carcinoma without lymph node metastasis. Estrogen and progesterone receptors were negative. Three years after operation without any adjuvant treatment, she has no recurrence of lesion.

[A Case of Metastatic Pancreatic Ductal Adenocarcinoma Successfully Treated with Modified FOLFIRINOX and Concurrent Radiotherapy].

A 70-year-old man with metastatic pancreatic ductal adenocarcinoma(cT4N1bM1, cStage Ⅳ)underwent chemotherapy with modified FOLFIRINOX without any severe adverse event to 20 cycles. In the middle of that, concurrent irradiation toward primary lesion(total dose, 43.2 Gy)was administered. Grade 1 adverse events include anemia, thrombocytopenia, hypoalbuminemia, hypokalemia, alkaline phosphatase increased, hypertension, peripheral sensory neuropathy, fatigue, anorexia and nausea. The relative dose intensities of oxaliplatin, irinotecan and fluorouracil at 6 months after beginning of treatment were 77.6, 84.0 and 88.3 percent, respectively. The total dose of administered oxaliplatin was 825 mg to the square meter. The primary lesion had been stable for the 20 cycles, although peritoneal dissemination had progressively increased in size. For 17 months, opioid was not necessary for the control of abdominal or back pain to the end of third-line treatment. Though safety or clinical benefits of modified FOLFIRINOX plus concurrent radiotherapy for metastatic pancreatic ductal adenocarcinoma have not been reported, in this case, such treatment might contribute to prolong prognosis or prevent developing abdominal or back pain.

[A Case of Malignant Thyroid Lymphoma That Was Difficult to Differentiate from Methotrexate-Associated Lymphoproliferative Disorders(MTX-LPD)].

A female patient in her 60s was going to get treatment for rheumatoid arthritis(RA). Considering the possibility of using biologics, CT examination was performed for screening of malignant diseases. A mass shadow in the left lobe of the thyroid gland was detected. The patient was followed up, and ultrasonography did not reveal any malignant findings. She was treated with methotrexate(MTX), and 1 year later, the thyroid mass was enlarged on CT. Ultrasonography revealed an enlarged hypoechoic region. Fine needle aspiration cytology revealed malignant lymphoma. Excisional biopsy was performed to determine the treatment plan. The pathological diagnosis was follicular lymphoma, and the possibility of methotrexate- associated lymphoproliferative disorders(MTX-LPD)persisted. It was difficult to discontinue MTX because of the high activity of RA. She was treated with rituximab for malignant lymphoma and concurrently with MTX for RA. The thyroid tumor disappeared for 3 months. Four years later, there is no sign of tumor recurrence.

Recurrent position and innervation pattern of recurrent peroneal nerve: A cadaveric study.

BACKGROUND: The recurrent peroneal nerve (RPN) branches from the common peroneal nerve or the deep peroneal nerve and it innervates to the lower patellar region. It has recently been reported that damage to the RPN causes pain in the lower patellar region; therefore, this study examined the recurrent position and the innervation pattern of the RPN. METHODS: Cases of knee deformity or atrophy were excluded, and 50 legs (25 males and 25 females) of 34 cadavers (15 males and 19 females) were examined to assess the recurrent position and the innervation pattern of the RPN. RESULTS: The recurrent position of the RPN was 27.9 ± 3.6 mm from the tip of the fibula. The RPN innervated to the patellar tendon in five of the 50 legs (10%), to the infrapatellar fat pad in 13 legs (26%), and to both the patellar tendon and the infrapatellar fat pad in 20 legs (40%), and to neither the patellar tendon nor the infrapatellar fat pad in 12 legs (24%). No significant sex differences were observed in the recurrent position and the innervation pattern of the RPN. CONCLUSIONS: In all cases, the recurrent position of the RPN was almost fixed from the tip of the fibula. The RPN frequently innervated to the patellar tendon or the infrapatellar fat pad (76%) in both males and females. These findings would be useful in knee surgery to preserve the RPN or for the diagnosis of pain in the lower patellar region.

[Two Cases of Stage â £ Occult Breast Cancer].

The first case is a 62-year-old female who complained of painful left axillary lymph node swelling. Six months later, a CT scan revealed multiple lung nodules. Biopsies of the axillary lymph node and lung showed metastatic carcinoma from breast cancer. However, no breast tumor was found. She was diagnosed with occult breast cancer with metastasis to the axillary lymph node and lung. ER(+), PgR(±), HER2(1+). Letrozole was administered, and effective control was achieved for 20 months. The second case is a 62-year-old female who presented with back pain. A CT scan revealed left axillary lymph node swelling and multiple osteolytic changes in the thoracolumbar spine and rib. Biopsies of the axillary lymph node and thoracic spine showed metastatic carcinoma from breast cancer. However, no breast tumor was found. She was diagnosed with occult breast cancer with metastasis to the axillary lymph nodule and bone. ER(+), PgR(+), HER2(1+). Fulvestrant and denosumab were administered. However, after 6 months, she discontinued the treatment. Our results suggested that effective control could be achieved through systemic therapy and local therapy was not necessary for Stage Ⅳ occult breast cancer.

[A Case of Breast Metastasis of Gastric Cancer after an Eight Year Latency Period].

A 66-year-old woman underwent distal gastrectomy because of gastric cancer(stage ⅠB)and received no adjuvant chemotherapy. Eight years after the operation, computed tomography showed a small nodule in the right breast. Mammography did not reveal any abnormalities. Ultrasound sonography showed a diffuse and gradual non-mass-like low echoic lesion. Core needle biopsy indicated a malignancy. Partial resection of the right breast was performed to obtain a diagnosis. On postoperative histopathological examination, signet-ring cells were found in the tumor, and immunohistochemical analysis showed that both the breast tumor and the gastric carcinoma were MUC5AC-positive and MUC1-negative. We diagnosed this breast tumor as metastasis from gastric cancer. The patient has received chemotherapy with no subsequent metastatic tumors, and good control has been achieved for 21 months after the detection of the breast metastasis.

Universal glass-forming behavior of in vitro and living cytoplasm.

Physiological processes in cells are performed efficiently without getting jammed although cytoplasm is highly crowded with various macromolecules. Elucidating the physical machinery is challenging because the interior of a cell is so complex and driven far from equilibrium by metabolic activities. Here, we studied the mechanics of in vitro and living cytoplasm using the particle-tracking and manipulation technique. The molecular crowding effect on cytoplasmic mechanics was selectively studied by preparing simple in vitro models of cytoplasm from which both the metabolism and cytoskeletons were removed. We obtained direct evidence of the cytoplasmic glass transition; a dramatic increase in viscosity upon crowding quantitatively conformed to the super-Arrhenius formula, which is typical for fragile colloidal suspensions close to jamming. Furthermore, the glass-forming behaviors were found to be universally conserved in all the cytoplasm samples that originated from different species and developmental stages; they showed the same tendency for diverging at the macromolecule concentrations relevant for living cells. Notably, such fragile behavior disappeared in metabolically active living cells whose viscosity showed a genuine Arrhenius increase as in typical strong glass formers. Being actively driven by metabolism, the living cytoplasm forms glass that is fundamentally different from that of its non-living counterpart.

High-resolution microrheology in the pericellular matrix of prostate cancer cells.

Many cells express a membrane-coupled external mechanical layer, the pericellular matrix (PCM), which often contains long-chain polymers. Its role and properties are not entirely known, but its functions are believed to include physical protection, mechanosensing, chemical signalling or lubrication. The viscoelastic response of the PCM, with polysaccharides as the main structural components, is therefore crucial for the understanding of its function. We have here applied microrheology, based on optically trapped micrometre-sized colloids, to the PCM of cultured PC3 prostate cancer cells. This technology allowed us to measure the extremely soft response of the PCM, with approximately 1 µm height resolution. Exogenously added aggrecan, a hyaluronan-binding proteoglycan, caused a remarkable increase in thickness of the viscoelastic layer and also triggered filopodia-like protrusions. The viscoelastic response of the PCM, however, did not change significantly.

(-)-Epigallocatechin-3-gallate suppresses growth of AZ521 human gastric cancer cells by targeting the DEAD-box RNA helicase p68.

(-)-Epigallocatechin-3-gallate (EGCG), the most abundant and biologically active polyphenol in green tea, induces apoptosis and suppresses proliferation of cancer cells by modulating multiple signal transduction pathways. However, the fundamental mechanisms responsible for these cancer-preventive effects have not been clearly elucidated. Recently, we found that EGCG can covalently bind to cysteine residues in proteins through autoxidation and subsequently modulate protein function. In this study, we demonstrate the direct binding of EGCG to cellular proteins in AZ521 human gastric cancer cells by redox-cycle staining. We comprehensively explored the binding targets of EGCG from EGCG-treated AZ521 cells by proteomics techniques combined with the boronate-affinity pull-down method. The DEAD-box RNA helicase p68, which is overexpressed in a variety of tumor cells and plays an important role in cancer development and progression, was identified as a novel EGCG-binding target. Exposure of AZ521 cells to EGCG lowered the p68 level dose dependently. The present findings show that EGCG inhibits AZ521 cell proliferation by preventing ß-catenin oncogenic signaling through proteasomal degradation of p68 and provide a new perspective on the molecular mechanism of EGCG action.

Generation of IFN-gamma-producing cells that recognize the major piroplasm surface protein in Theileria orientalis-infected bovines.

Theileriosis is a tick-borne protozoan disease caused by Theileria species. The Theileria species are classified into two groups depending on the cell type in which they proliferate and the clinical symptoms. The first group consists of lymphoproliferative Theileria species (T. parva and T. annulata), which mainly proliferate in lymphocytes, causing uncontrolled lymphocyte proliferation. The other group consists of a nonlymphoproliferative Theileria species (T. orientalis, also known as T. sergenti) that proliferates in erythrocytes and causes hemolytic anemia. Based on reports of generation of antigen-specific CD4(+) and CD8(+) T cells in lymphoproliferative theileriosis, we investigated whether T cells specific to the T. orientalis antigen are present in the nonlymphoproliferative form of the disease. In this study, we developed a new assay based on an enzyme-linked immunospot (ELISpot) to detect interferon-gamma (IFN-gamma)- and interleukin-10 (IL-10)-secreting cells in a series of cryogenically preserved bovine peripheral blood mononuclear cells (PBMCs). We first determined that IFN-gamma- and IL-10-secreting T cells were present in PBMCs by stimulating them with phytohemagglutinin L (PHA-L=red kidney bean lectin L, known as T cell stimulator), and then determined whether T. orientalis-specific T cells are present in T. orientalis-infected bovines. Peptides derived from T. orientalis major piroplasm surface protein (MPSP) were used as a T. orientalis-specific stimulator in the ELISpot assay, and peptides from glycoprotein B (gB) of the bovine herpes virus-1 (BHV-1) were used as a BHV-1-specific stimulator as a control for monitoring the immune response. Compared with results obtained using the BHV-1 (gB peptides)-specific IFN-gamma ELISpot assay to assess BHV-1-immunized Holsteins, prominent T. orientalis MPSP peptide-specific IFN-gamma and IL-10 positive spots were detected in T. orientalis-infected Holsteins but weak positive responses were exhibited by T. orientalis-infected Angus and Japanese Black cattle. As far as we are aware, this is the first report to show direct evidence of the presence of T. orientalis-specific T cells in T. orientalis-infected bovines using an antigen-specific ELISpot assay system and that T. orientalis-specific, IFN-gamma- and IL-10-producing T cells are produced in T. orientalis-infected Holsteins.

Covalent modification of proteins by green tea polyphenol (-)-epigallocatechin-3-gallate through autoxidation.

Green tea polyphenol (-)-epigallocatechin-3-gallate (EGCG) has various beneficial properties including chemopreventive, anticarcinogenic, and antioxidant actions. The interaction with proteins known as EGCG-binding targets may be related to the anticancer effects. However, the binding mechanisms for this activity remain poorly understood. Using mass spectrometry and chemical detection methods, we found that EGCG forms covalent adducts with cysteinyl thiol residues in proteins through autoxidation. To investigate the functional modulation caused by binding of EGCG, we examined the interaction between EGCG and a thiol enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Concentration-dependent covalent binding of EGCG to GAPDH was found to be coupled to the irreversible inhibition of GAPDH activity. Mutation experiments revealed that EGCG is primarily bound to the cysteinyl thiol group of the active center, indicating that the irreversible inhibition of GAPDH is due to the covalent attachment of EGCG to the active-center cysteine. Moreover, using EGCG-treated cancer cells, we identified GAPDH as a target of EGCG covalent binding through specific interactions between catechols and aminophenyl boronate agarose resin. Based on these findings, we propose that the covalent modification of proteins by EGCG may be a novel pathway related to the biological activity of EGCG.

Effect of posterior sub-tenon administration of triamcinolone acetonide on leukocyte dynamics in rat retinal microcirculation after panretinal photocoagulation.

PURPOSE: Macular edema is one of the serious side effects associated with panretinal photocoagulation (PRP). The inhibitory effect of triamcinolone acetonide (TA) on leukocyte-endothelial cell interactions in vivo after PRP was evaluated. METHODS: Argon laser photocoagulation was performed in one half of the retinas in male Brown Norway rats. Experimental rats were injected with 2 mg TA (50-microL volume) in the posterior sub-Tenon space, and the vehicle-treated rats were injected with the same amount of saline (50 microL) immediately after PRP. Untreated rats were used as the control. Leukocyte dynamics in retinal microcirculation and retinal vessel diameters were evaluated 1 day after laser photocoagulation with the use of acridine orange digital fluorography. Retinal thickness was evaluated with optical coherence tomography. RESULTS: The number of rolling leukocytes and accumulating leukocytes in the retina decreased by 66% in the TA-treated rats (P < 0.01) and by 24% (P < 0.05), respectively, compared with the number in the vehicle-treated rats. Retinal thickness in the vehicle-treated rats was significantly thicker than that in control rats 1 day after laser photocoagulation (P < 0.01). Retinal thickness in the TA-treated rats was significantly suppressed compared with that in the vehicle-treated rats (P < 0.05). CONCLUSIONS: Sub-Tenon administration of TA significantly suppressed leukocyte dynamics in rat retinal microcirculation and decreased retinal edema after laser photocoagulation. The results suggest that the suppression of leukocyte-endothelial cell interactions in retinal microcirculation may be one mechanism responsible for the therapeutic effect of sub-Tenon TA on postlaser retinal edema.

Nonequilibrium mechanics of active cytoskeletal networks.

Cells both actively generate and sensitively react to forces through their mechanical framework, the cytoskeleton, which is a nonequilibrium composite material including polymers and motor proteins. We measured the dynamics and mechanical properties of a simple three-component model system consisting of myosin II, actin filaments, and cross-linkers. In this system, stresses arising from motor activity controlled the cytoskeletal network mechanics, increasing stiffness by a factor of nearly 100 and qualitatively changing the viscoelastic response of the network in an adenosine triphosphate-dependent manner. We present a quantitative theoretical model connecting the large-scale properties of this active gel to molecular force generation.